Advertisement

Isolation and characterization of hexokinase genes PsHXK1 and PsHXK2 from tree peony (Paeonia suffruticosa Andrews)

  • Chao Zhang
  • Lili Zhang
  • Jianxin Fu
  • Li DongEmail author
Original Article
  • 49 Downloads

Abstract

Hexokinase (HXK) plays important roles in hexose phosphorylation and sugar signaling. HXK regulates the glucose-induced accumulation of anthocyanin in many species. Little is known about the biological function of the HXK gene family in Paeonia suffruticosa. cDNA sequences of two hexokinase genes PsHXK1 and PsHXK2 were isolated using RACE-PCR and RT-PCR from P. suffruticosa. PsHXK1 encodes 498 amino acids with a 1497-bp open reading frame (ORF), and PsHXK2 contains 493 amino acids with a 1482-bp ORF. Sequence and phylogenetic analyses suggest that PsHXK1 and PsHXK2 belong to type-B HXK and may function as glucose sensors. PsHXK1 and PsHXK2 mRNA were detected in all tested tissues. PsHXK1 is highly expressed in petals and stamens, while PsHXK2 is highly expressed in stamens. At the former stages of flower opening, PsHXK1 and PsHXK2 show higher expression levels in on-tree flowers compared with cut flowers. Overexpressing PsHXK1 and PsHXK2 in Arabidopsis enhances glucose sensitivity, inhibits plant growth in response to glucose, and induces anthocyanin accumulation in response to the high level of glucose. Overall, our results primarily reveal the biological function of PsHXK1 and PsHXK2, especially their involvement in glucose-induced anthocyanin accumulation.

Keywords

Paeonia suffruticosa Hexokinase Anthocyanin accumulation Glucose sensor 

Notes

Acknowledgements

This work was supported by the National Natural Science Foundation of China (31572164 and 30972030) and National Key Research and Development Project (2018YFD1000407).

Author contributions

LD conceived and designed the research. CZ, LZ and JF conducted the experiments. CZ analyzed the data. CZ and LD wrote the manuscript. All the authors read and approved the manuscript.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

This article does not contain any studies conducted on human or animal subjects.

Supplementary material

11033_2019_5135_MOESM1_ESM.jpg (553 kb)
Supplementary material 1 Fig. S1 PCR amplification of isolation of PsHXK1 (A) and PsHXK2 (B) in P. suffruticosa. M: DL2000 marker; M′: DL2000 plus marker; Lane 1: PCR amplification of 3′ end of cDNA sequence of PsHXK1; Lane 2: PCR amplification of 5′ end of cDNA sequence of PsHXK1; Lane 3: PCR amplification of ORF sequence of PsHXK1; Lane 4: PCR amplification of 3′ end of cDNA sequence of PsHXK2; Lane 5: PCR amplification of 5′ end of cDNA sequence of PsHXK2; Lane 6: PCR amplification of ORF sequence of PsHXK2 (JPEG 552 kb)
11033_2019_5135_MOESM2_ESM.jpg (130 kb)
Supplementary material 2 Fig. S2 Comparison of the N-terminal region of PsHXK1 and PsHXK2 proteins with other plant HXK proteins. TargetP scores and predictions are shown on the right. cTP: chloroplast transit peptide; mTP: mitochondrial targeting peptide; SP: secretory pathway signal peptide; other: any other location; S: predicted secretory pathway. For each protein, the predicted location with the highest score is shown in bold. Predicted membrane anchor of PsHXK1 and PsHXK2 is annotated in the box (JPEG 129 kb)
11033_2019_5135_MOESM3_ESM.tif (626 kb)
Supplementary material 3 Fig. S3 Expression of the endogenous hexokinase genes AtHXK1, AtHXK2, AtHXK3, AtHKL1 and AtHKL2 in transgenic Arabidopsis overexpressing PsHXK1 or PsHXK2 and wild-type (WT) Arabidopsis using semiquantitative RT-PCR. AtUBQ5 was used as an internal reference gene (TIFF 625 kb)
11033_2019_5135_MOESM4_ESM.docx (15 kb)
Supplementary material 4 (DOCX 14 kb)

References

  1. 1.
    Karve A, Rauh BL, Xia X, Kandasamy M, Meagher RB, Sheen J, Moore B (2008) Expression and evolutionary features of the hexokinase gene family in Arabidopsis. Planta 228:411–425.  https://doi.org/10.1007/s00425-008-0746-9 CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Yanagisawa S, Yoo SD, Sheen J (2003) Differential regulation of EIN3 stability by glucose and ethylene signalling in plants. Nature 425:521–525.  https://doi.org/10.1038/nature01984 CrossRefPubMedGoogle Scholar
  3. 3.
    Moore B, Zhou L, Rolland F, Hall Q, Cheng WH, Liu YX, Hwang I, Jones T, Sheen J (2003) Role of the arabidopsis glucose sensor HXK1 in nutrient, light, and hormonal signaling. Science 300:332–336.  https://doi.org/10.1126/science.1080585 CrossRefPubMedGoogle Scholar
  4. 4.
    Cho J-I, Ryoo N, Eom J-S, Lee D-W, Kim H-B, Jeong S-W, Lee Y-H, Kwon Y-K, Cho M-H, Bhoo S-H, Hahn T-R, Park Y-I, Hwang I, Sheen J, Jeon J-S (2009) Role of the rice hexokinases OsHXK5 and OsHXK6 as glucose sensors. Plant Physiol 149:745–759.  https://doi.org/10.1104/pp.108.131227 CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Kandel-Kfir M, Damari-Weissler H, German MA, Gidoni D, Mett A, Belausov E, Petreikov M, Adir N, Granot D (2006) Two newly identified membrane-associated and plastidic tomato HXKs: characteristics, predicted structure and intracellular localization. Planta 224:1341–1352.  https://doi.org/10.1007/s00425-006-0318-9 CrossRefPubMedGoogle Scholar
  6. 6.
    Olsson T, Thelander M, Ronne H (2003) A novel type of chloroplast stromal hexokinase is the major glucose-phosphorylating enzyme in the moss Physcomitrella patens. J Biol Chem 278:44439–44447.  https://doi.org/10.1074/jbc.M306265200 CrossRefPubMedGoogle Scholar
  7. 7.
    Kim M, Lim JH, Ahn CS, Park K, Kim GT, Kim WT, Pai HS (2006) Mitochondria-associated hexokinases play a role in the control of programmed cell death in Nicotiana benthamiana. Plant Cell 18:2341–2355.  https://doi.org/10.1105/tpc.106.041509 CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Kelly G, Sade N, Attia Z, Secchi F, Zwieniecki M, Holbrook NM, Levi A, Alchanatis V, Moshelion M, Granot D (2014) Relationship between hexokinase and the aquaporin PIP1 in the regulation of photosynthesis and plant growth. PLoS ONE 9:e87888.  https://doi.org/10.1371/journal.pone.0087888 CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Sarowar S, Lee J-Y, Ahn E-R, Pai H-S (2008) A role of hexokinases in plant resistance to oxidative stress and pathogen infection. J Plant Biol 51:341–346.  https://doi.org/10.1007/BF03036136 CrossRefGoogle Scholar
  10. 10.
    Kim Y-M, Heinzel N, Giese JO, Koeber J, Melzer M, Rutten T, Von Wiren N, Sonnewald U, Hajirezae M-R (2013) A dual role of tobacco hexokinase 1 in primary metabolism and sugar sensing. Plant Cell Environ 36:1311–1327.  https://doi.org/10.1111/pce.12060 CrossRefPubMedGoogle Scholar
  11. 11.
    Hu D-G, Sun C-H, Zhang Q-Y, An J-P, You C-X, Hao Y-J (2016) Glucose sensor MdHXK1 phosphorylates and stabilizes MdbHLH3 to promote anthocyanin biosynthesis in apple. PLoS Genet 12:e1006273.  https://doi.org/10.1371/journal.pgen.1006273 CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Zhang C, Wang W, Wang Y, Gao S, Du D, Fu J, Dong L (2014) Anthocyanin biosynthesis and accumulation in developing flowers of tree peony (Paeonia suffruticosa) ‘Luoyang Hong’. Postharvest Biol Technol 97:11–22.  https://doi.org/10.1016/j.postharvbio.2014.05.019 CrossRefGoogle Scholar
  13. 13.
    Zhang C, Fu J, Wang Y, Gao S, Du D, Fan W, Jia G, Li D (2015) Glucose supply improves petal coloration and anthocyanin biosynthesis in Paeonia suffruticosa ‘Luoyang Hong’ cut flowers. Postharvest Biol Technol 101:73–81.  https://doi.org/10.1016/j.postharvbio.2014.11.009 CrossRefGoogle Scholar
  14. 14.
    Moalem-Beno D, Tamari G, Leitner-Dagan Y, Borochov A, Weiss D (1997) Sugar-dependent gibberellin-induced chalcone synthase gene expression in petunia corollas. Plant Physiol 113:419–424.  https://doi.org/10.1104/pp.113.2.419 CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Neta-Sharir I, Shoseyov O, Weiss D (2000) Sugars enhance the expression of gibberellin-induced genes in developing petunia flowers. Physiol Plant 109:196–202.  https://doi.org/10.1034/j.1399-3054.2000.100212.x CrossRefGoogle Scholar
  16. 16.
    Zheng Y, Tian L, Liu H, Pan Q, Zhan J, Huang W (2009) Sugars induce anthocyanin accumulation and flavanone 3-hydroxylase expression in grape berries. Plant Growth Regul 58:251–260.  https://doi.org/10.1007/s10725-009-9373-0 CrossRefGoogle Scholar
  17. 17.
    Vitrac X, Larronde F, Krisa S, Decendit A, Deffieux G, Mérillon J-M (2000) Sugar sensing and Ca2+-calmodulin requirement in Vitis vinifera cells producing anthocyanins. Phytochemistry 53:659–665.  https://doi.org/10.1016/S0031-9422(99)00620-2 CrossRefPubMedPubMedCentralGoogle Scholar
  18. 18.
    Chang S, Puryear J, Cairney J (1993) A simple and efficient method for isolating RNA from pine trees. Plant Mol Biol Rep 11:113–116.  https://doi.org/10.1007/BF02670468 CrossRefGoogle Scholar
  19. 19.
    Zhang C, Wang Y, Fu J, Dong L, Gao S, Du D (2014) Transcriptomic analysis of cut tree peony with glucose supply using the RNA-Seq technique. Plant Cell Rep 33:111–129.  https://doi.org/10.1007/s00299-013-1516-0 CrossRefPubMedGoogle Scholar
  20. 20.
    Wang YJ, Dong L, Zhang C, Wang XQ (2012) Reference gene gelection for real-time quantitative PCR normalization in tree peony (Paeonia suffruticosa Andr.). J Agric Biotechnol 20:521–528Google Scholar
  21. 21.
    Wang J, Han K, Dai S (2009) Construction of expression vector and transformation of chrysanthemum with maize Lc gene. Genom Appl Biol 28:229–236Google Scholar
  22. 22.
    Clough SJ, Bent AF (1998) Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. Plant J 16:735–743.  https://doi.org/10.1046/j.1365-313x.1998.00343.x CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Jeong S-W, Das PK, Jeoung SC, Song J-Y, Lee HK, Kim Y-K, Kim WJ, Park YI, Yoo S-D, Choi S-B (2010) Ethylene suppression of sugar-induced anthocyanin pigmentation in Arabidopsis. Plant Physiol 154:1514–1531.  https://doi.org/10.1104/pp.110.161869 CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Cho J-I, Ryoo N, Ko S, Lee S-K, Lee J, Jung K-H, Lee Y-H, Bhoo SH, Winderickx J, An G, Hahn T-R, Jeon J-S (2006) Structure, expression, and functional analysis of the hexokinase gene family in rice (Oryza sativa L.). Planta 224:598–611.  https://doi.org/10.1007/s00425-006-0251-y CrossRefPubMedGoogle Scholar
  25. 25.
    Wang J, Wang X, Geng S, Singh SK, Wang Y, Pattanaik S, Yuan L (2018) Genome-wide identification of hexokinase gene family in Brassica napus: structure, phylogenetic analysis, expression, and functional characterization. Planta 248:171–182.  https://doi.org/10.1007/s00425-018-2888-8 CrossRefPubMedGoogle Scholar
  26. 26.
    Karve R, Lauria M, Virnig A, Xia X, Rauh BL, Moore BD (2010) Evolutionary lineages and functional diversification of plant hexokinases. Mol Plant 3:334–346.  https://doi.org/10.1093/mp/ssq003 CrossRefPubMedGoogle Scholar
  27. 27.
    Cho Y-H, Yoo S-D, Sheen J (2006) Regulatory functions of nuclear hexokinase1 complex in glucose signaling. Cell 127:579–589.  https://doi.org/10.1016/j.cell.2006.09.028 CrossRefPubMedGoogle Scholar
  28. 28.
    Wang X, Li L, Yang P, Gong C (2014) The role of hexokinases from grape berries (Vitis vinifera L.) in regulating the expression of cell wall invertase and sucrose synthase genes. Plant Cell Rep 33:337–347.  https://doi.org/10.1007/s00299-013-1533-z CrossRefPubMedGoogle Scholar
  29. 29.
    Price J, Laxmi A, Martin SKS, Jang J-C (2004) Global transcription profiling reveals multiple sugar signal transduction mechanisms in Arabidopsis. Plant Cell 16:2128–2150.  https://doi.org/10.1105/tpc.104.022616 CrossRefPubMedPubMedCentralGoogle Scholar
  30. 30.
    Shi G, Guo X, Zhang G, Bao M (2009) Analysis of sugar metabolism during florescence and flower senescence of tree peony petal. Acta Hortic Sinica 36:1184–1190Google Scholar
  31. 31.
    Jang JC, Leon P, Zhou L, Sheen J (1997) Hexokinase as a sugar sensor in higher plants. Plant Cell 9:5–19.  https://doi.org/10.1105/tpc.9.1.5 CrossRefPubMedPubMedCentralGoogle Scholar
  32. 32.
    Hara M, Oki K, Hoshino K, Kuboi T (2003) Enhancement of anthocyanin biosynthesis by sugar in radish (Raphanus sativus) hypocotyls. Plant Sci 164:259–265.  https://doi.org/10.1016/S0168-9452(02)00408-9 CrossRefGoogle Scholar
  33. 33.
    Solfanelli C, Poggi A, Loreti E, Alpi A, Perata P (2006) Sucrose-specific induction of the anthocyanin biosynthetic pathway in Arabidopsis. Plant Physiol 140:637–646.  https://doi.org/10.1104/pp.105.072579 CrossRefPubMedPubMedCentralGoogle Scholar
  34. 34.
    Kunz S, Gardeström P, Pesquet E, Kleczkowski LA (2015) Hexokinase 1 is required for glucose-induced repression of bZIP63, At5g22920, and BT2 in Arabidopsis. Front Plant Sci 6:525.  https://doi.org/10.3389/fpls.2015.00525 CrossRefPubMedPubMedCentralGoogle Scholar
  35. 35.
    Kunz S, Pesquet E, Kleczkowski LA (2014) Functional dissection of sugar signals affecting gene expression in Arabidopsis thaliana. PLoS ONE 9(6):e100312.  https://doi.org/10.1371/journal.pone.0100312 CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  1. 1.Beijing Key Laboratory of Ornamental Plants Germplasm Innovation and Molecular Breeding, National Engineering Research Center for Floriculture and College of Landscape ArchitectureBeijing Forestry UniversityBeijingChina
  2. 2.Department of Ornamental Horticulture, School of Landscape ArchitectureZhejiang Agriculture and Forestry UniversityHangzhouChina

Personalised recommendations